Project Summary Newborns and young infants are at risk of infection and often respond sub-optimally to vaccines. HIPC Project 3 proposes to leverage two technological advances that will provide fundamental new insights into vaccine- induced molecular changes associated with protective immunity and may transform future neonatal vaccine development: 1) novel ?OMIC? technologies capable of measuring comprehensive molecular signatures, in particular transcriptomics and proteomics, following immunization; and 2) the development of human in vitro systems to model age-specific innate and adaptive immune responses to immunization. Our preliminary data suggest that vaccines currently given at birth, the Alum-adjuvanted hepatitis B vaccine (HBV) and the live attenuated Mycobacterium bovis named Bacille Calmette-Gurin (BCG), induce age-dependent immune and OMIC responses, and that these distinct in vivo-induced changes can also be modeled in vitro. Accordingly, the goal of Project 3 is to characterize age-specific OMIC responses to immunization in vitro and in vivo in well- defined cohorts to deepen our understanding of vaccine-induced signatures that predict vaccine immunogenicity, including correlates of protection (CoP), and enhance future age-specific vaccine development. We will achieve this goal by pursuing the following Specific Aims: in Specific Aim 1, we will characterize vaccine-induced molecular pathways correlating with CoP in vivo employing whole blood assays in vitro; and in Specific Aim 2, we will characterize vaccine-induced molecular pathways correlating with CoP in vivo employing human microphysiologic tissue constructs in vitro HIPC Project 3 will work in synergy with the other HIPC components, including Projects-1 (OMIC integration) and -2 (Immunophenotyping) and the Data Management Core to define and validate signatures that correlate with neonatal vaccine immunogenicity. These studies will leverage novel in vitro platforms to define, characterize and dissect molecular signatures corresponding to newborn vaccine immunogenicity, thereby informing future early life vaccine development. !